Evaluation of methods for the determination of mitochondrial respiratory chain enzyme activities in human skeletal muscle samples

The quantification of mitochondrial enzyme activities in skeletal muscle sa mples of patients suspected of having mitochondrial myopathies is problemat ic. Therefore, we have evaluated different methods for the determination of activities cytochrome c oxidase and NADH:CoQ oxidoreductase in human skele tal muscle samples. The measurement of cytochrome c oxidase activity in the presence of 200 mu M ferrocytochrome c and the detection of NADH:CoQ oxido reductase as rotenone-sensitive NADH:CoQ(1) reductase resulted in comparabl e citrate synthase-normalized respiratory chain enzyme activities of both i solated mitochondria and homogenates from control human skeletal muscle sam ples. These methods allowed the precise detection of deficiencies of respir atory chain enzymes in skeletal muscle of two patients harboring only 20 an d 27% of deleted mitochondrial DNA, respectively. Therefore, citrate syntha se-normalized respiratory chain activities can serve as stable reference va lues for the determination of a putative mitochondrial defect in human skel etal muscle. (C) 2000 Academic Press.